A binary self-assembled monolayer (SAM) of 11-mercaptoundecarmit acid (MUA) arid 3,6-clioxa-8-mercaptooctan-1-ol (DMOL) was formed on a gold electrode as a base for protein aptamer immobilization. The aptamer, specific to hemoglobin, was bound to MUA. A DMOL SAM was used to prevent nonspecific protein adsorption. Electrochemical impedance spectroscopy was performed to determine the electrOchemical dhacteristic of a unary SAM of MUA and a binary SAM consisting of MUA and DMOL. The DMOL SAM preVented adsorption of nonspecific proteins (e.g., thrombin). For the binary SAM, 20:80, 5050, and 80:20 mol MUA and DMOL were used. The antifouling effect against nonspecific proteins was enhanced as-the ratio of -DMOL increased, but the signal intensity (impedance change) decreased. The binary SAM with 50:50 mol % MUA and DMOL showed repelling of nonspecific adsorption of thrombin (70.5% decrease in the impedance change), and it still generated a sufficient signal for target molecules. Additional tests with..pure protein samples showed that the presence of DMOL did not affect the aptamer function.